Anti-acne and Tyrosinase Inhibition Properties of Taxifolin and Some Flavanonol Rhamnosides from Kempas (Koompassia malaccensis)

Irmanida Batubara, Harlinda Kuspradini, and Tohru Mitsunaga

Abstract

Taxifolin (1) and some flavanonol rhamnosides (neoastilbin (2), astilbin (3), and isoastilbin (4)) have been isolated from kempas (Koompassia malaccensis). Our previous research about antimicrobial activity against Streptococcus sobrinus and glucosyltransferase inhibitory activity of these compounds have been reported. Now, we carried out the anti- acne and tyrosinase inhibition properties of all four compounds. Antimicrobial against Propionibacterium acnes, P. acnes lipase inhibitory activity and antioxidant activity were established for anti-acne activity. Tyrosinase inhibition property was measured using L-tyrosine and L-DOPA as substrate. The results for anti-acne showed that no antimicrobial activity against P. acnes for all compounds, the best lipase inhibition properties showed on compound 4 with IC50 about 1.36 mg/ml, and % inhibition for antioxidant at concentration 10 mg/ml are 31.16, 25.64, 28.47, and 31.01% respectively. Tyrosinase inhibition of compound 1 at concentration 1 mg/ml is 24.12% for monophenolase and 5.18% for diphenolase. Compound 2 has tyrosinase inhibition about 25.95% (monophenolase) and 14.18% (diphenolase) at concentration 1 mg/ml. Compound 3 has tyrosinase inhibition about 27.17% (monophenolase) and 6.23% (diphenolase) at same concentration, while compound 4 has tyrosinase inhibition about 11.17% (monophenolase) and 9.75% (diphenolase).

Key words: taxifolin, flavanonol rhamnosides, anti-acne, tyrosinase inhibition, Koompassia malaccensis.

Introduction enzyme may lead to development of novel skin whitening agents. Acne is a very common skin disease characterized Kempas (Koompassia malaccensis) is an important by pimples on the face, chest, and back. It occurs when and valuable wood because it is used as flooring, the pores of the skin become clogged with oil, dead skin moldings, furniture, and veneer in Indonesia. It belongs to cells, and bacteria. Acne is not a simple disease; it may the Leguminosae family and widely distributed in sometimes lead to social phobia, lowered self-image, and Sumatera and Kalimantan. Many plants in Leguminosae depression (Koo and Smith 1991). show bioactivities such as antidiabetic, antimalaria, and Compounds targeting acne therefore should be able antioxidant. Traditionally, the bark of kempas is used to to inhibit P. acnes population and inhibit P. acnes lipase prepare medicinal baths because of its antifever and activity, as a result reduce pro-inflammatory lipids in antidysentery activities (Kobayashi et al. 1996). sebum as well as reduce post-acne scar formation. The In previous report (Kuspradini et al. 2009), taxifolin materials that have antioxidant activity may be useful for (1) and some flavanonol rhamnosides (neoastilbin (2), relieving hypertrophic scars and keloid formation on the astilbin (3), and isoastilbin (4) have been isolated from skin (Furakawa et al. 1995). In other words, compounds kempas (K. malaccensis). Some activities of taxifolin have or materials claiming good for acne control should been reported but not many activities report for flavanonol possess anti-bacterial, anti-lipase, anti-inflammatory, and rhamnosides. On this paper we performed antimicrobial antioxidant activities. activity against P. acnes, P. acnes lipase inhibitory Tyrosinase inhibitors have been a great concern activity, antioxidant activity and tyrosinase inhibitory solely due to the key role of tyrosinase in both mammalian activity of taxifolin and some flavanonol rhamnosides melanogenesis and fruit or fungi enzymatic browning isolated from kempas. (Chang 2009). Melanogenesis is a principal parameter of differentiation of melanocytes and melanoma cells Materials and Methods (Ohguchi et al. 2005). The formation of melanin in human body influenced or reduced by several mechanisms, Plant Material including anti-oxidation, direct tyrosinase inhibition, K. malaccensis was provided by the Department of melanin inhibition of migration from cell to cell and Forest Product Technology, Mulawarman University, hormonal activities etc (Pawelek and Korner 1982). Kalimantan. Its voucher specimen (FHT.LA.13.11m) was Tyrosinase is responsible for pigmentation of skin, eyes deposited at the Wood Anatomy Laboratory of and hair. It made tyrosinase inhibitors have been used Mulawarman University, Indonesia. frequently in cosmetics and depigmenting agents for hyperpigmentation. Investigation of inhibitors of this

Anti-acne and Tyrosinase Inhibition Properties of Taxifolin and Some Flavanonol Rhamnosides 45 from Kempas (Koompassia malaccensis) Irmanida Batubara, Harlinda Kuspradini, and Tohru Mitsunaga Fractionation, Isolation, and Identification of and 4. Additionally, a glycoside signals were observed at Compounds 1, 2, 3, and 4. 4.02~5.40 ppm, 3.50~4.16 ppm, 3.32~3.65 ppm, The fractionation, isolation and identification of 3.17~3.57 ppm, 2.27~4.22 ppm, and 0.88~1.15 ppm, compounds 1, 2, 3, and 4 were performed the same with which indicates rhamnose residue. Furthermore, the our previous report (Kuspradini et al. 2009). heteronuclear multiple bond coherence (HMBC) of these compounds showed a correlation between the anomeric Anti-acne Activity Assay proton of rhamnose and C-3 carbon of flavanonol, which Anti-acne activity assay was performed based indicates that rhamnose should be connected with C-3 of antimicrobial against P. acnes, P. acnes lipase inhibitory taxifolin moiety (Figure 2). activity, and antioxidant activity. All of these activities Therefore Compounds 2, 3, and 4 are assumed as were performed like methods in Batubara et al. (2009). flavanonol-3-O-rhamnoside. As the result of comparing the 1H NMR data of references Compound B, C, and D Tyrosinase Inhibitory Activity were identified as neoastilbin (De Britto et al. 1995), Inhibition of tyrosinase activity (monophenolase) and astilbin (Guo et al. 2007), and isoastilbin (Du et al. 2005), DOPA auto-oxidation (diphenolase). This assay was respectively, as illustrated in Figure 2. performed using methods as described earlier (Curto et al. The anti-acne properties of the 4 compounds start 1999; Nerya et al. 2003; Batubara et al. 2010). with antimicrobial test against P. acnes. The results showed that all compounds have no antimicrobial Results and Discussion properties. This result is not the same with our finding before that all of these compounds have antimicrobial Figure 1 shows HPLC of crude extract of Kempas. It activity against S. sobrinus (Kuspradini et al. 2009). was purified by p-HPLC to isolate the four compounds, Other anti-acne property tested is P. acnes lipase and identified by NMR. The UV-Vis spectra of some major inhibition activity. The result of this test is shown in Figure peaks, 1, 2, 3, and 4 were similar to that of taxifolin. By 3. Inhibition properties for lipase of compound 1, 2, 3, and analysis of the NMR data that was generated in this study, 4 at concentration 31.25 mg/ml are 39.18, 0.31, 39.24, and by comparison of the physical and spectral data with and 59.19% respectively. Since compound 4 is the best those reported in the literature, Compound 1 was compound for lipase inhibitor, IC50 value of compound 4, identified as taxifolin (Lee et al. 2003). isoastilbin was calculated. It showed that isoastilbin has 13 The C-NMR data of Compound 1 exhibited a typical IC50 about 1.36 mg/ml. This IC50 value is lower than IC50 signal of flavonol-type skeleton: 83 ppm at C-2 and 77 value of IPMP as positive control (166.4 mg/ml). The last ppm at C-3. The saturated bond between C-2 and C-3 activity for anti-acne is antioxidant property. The % was confirmed by the presence of doublets at 4.90 ppm inhibition for antioxidant of all compounds at concentration (H-2) and 4.49 ppm (H-3) in 1H-NMR spectrum. Thus, the 10 mg/ml are 31.16, 25.64, 28.47, and 31,01% compound 1 was identified as a dihydroquercetin/taxifolin. respectively (Figure 4). The flavanonol like taxifolin signals were commonly observed in 1H-NMR spectroscopies of Compound 2, 3,

Figure 1. HPLC chromatogram of Kempas crude extract. Column : VP-ODS (250mm x 4.6mm i.d.); gradient program : MeOH : 0.05%TFA = 10% : 90% - 80%: 20% (40 min), 100%:0% (50 min), wavelength : 280 nm; flow rate : 1 ml/min; analysis time: 50 min.

46 Wood Research Journal Vol.1 • No. 1 • 2010

Figure 2. Structures of taxifolin and flavanonol rhamnosides isolated from Koompassia malaccensis wood 50% ethanol extracts. Compound 1, 2, 3, and 4. a) compound 1 (taxifolin), b) 2 (neoastilbin), c) 3 (astilbin), d) 4 (isoastilbin).

Anti-acne and Tyrosinase Inhibition Properties of Taxifolin and Some Flavanonol Rhamnosides 47 from Kempas (Koompassia malaccensis) Irmanida Batubara, Harlinda Kuspradini, and Tohru Mitsunaga 70 60 50 40 30

% inihibition 20 10 0 1 2 3 4 compound

Figure 3. Lipase inhibitory activity of all compounds at concentration 31.25 mg/ml.

35 30 25 20 15

% inihibition 10 5 0 1 2 3 4 compound

Figure 4. Antioxidant activity of all compounds at concentration 10 mg/ml.

30 25 20 monophenolase 15 diphenolase 10 % inihibition 5 0 1 2 3 4 compound

Figure 5. Tyrosinase inhibition activity of all compounds at concentration 1mg/ml.

Tyrosinase activity of all compounds are not good. It Conclusions only showed that tyrosinase inhibition of compound 1 at concentration 1 mg/ml is 24.12% for monophenolase and The results for anti-acne showed that no 5.18% for diphenolase. Compound 2 has tyrosinase antimicrobial activity against P. acnes for all compounds. inhibition about 25.95% (monophenolase) and 14.18% The tyrosinase activities are also not good from all (diphenolase) at concentration 1 mg/ml. Compound 3 has compounds. tyrosinase inhibition about 27.17% (monophenolase) and Only isoastilbin showed the best lipase inhibition 6.23% (diphenolase) at same concentration, while properties with IC50 about 1.36 mg/ml, and % inhibition for compound 4 has tyrosinase inhibition about 11.17% antioxidant of taxifolin, neoastilbin, astilbin, and isoastilbin (monophenolase) and 9.75% (diphenolase). at concentration 10 mg/ml are 31.16, 25.64, 28.47, and 31.01% respectively.

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Anti-acne and Tyrosinase Inhibition Properties of Taxifolin and Some Flavanonol Rhamnosides 49 from Kempas (Koompassia malaccensis) Irmanida Batubara, Harlinda Kuspradini, and Tohru Mitsunaga